Institute of Animal Husbandry and Veterinary Medicine, Hubei Academy of Agricultural sciences, Wuhan, 430064, China.
In Vitro Cell Dev Biol Anim. 2019 Oct;55(9):733-740. doi: 10.1007/s11626-019-00388-7. Epub 2019 Aug 5.
Intestinal epithelial cells (IECs) not only have an absorption function but also act as a physical barrier between the body and the intestinal bacterial flora. Damage to IECs leads to the breakdown of this barrier and has negative effects on animal health. Intestinal epithelial damage is frequently associated with long-term acute stress, such as increased temperature and new stress management models. The intestinal epithelial damage caused by environmental stress has been linked to oxidative stress. Until now, the effects of intestinal epithelial antioxidant activity from feed additives and treatments could be tested in ducks only in vivo because of the lack of in vitro cell culture systems. In this study, we describe our protocol for the easy isolation and culture of IECs from the small intestine of duck embryos. Immunofluorescence was used for the cytological identification of IECs. In addition, IEC marker genes (IAP and CDH1) could also be detected in cultured cells. And cell status assessments were performed, and cell proliferation viability was analyzed by CCK-8 assay. Furthermore, we constructed an oxidative stress model to be used to research the oxidative stress response mechanism, and drugs acting on the cell signal transduction pathway. In conclusion, we have developed an effective and rapid protocol for obtaining duck primary IECs and constructed an oxidative stress model. These IECs exhibit features consistent with epithelial cells and could be used to explore the physiological mechanisms of oxidative stress ex vivo.
肠上皮细胞(IECs)不仅具有吸收功能,而且在机体与肠道细菌菌群之间起到物理屏障的作用。IECs 的损伤会导致该屏障的破坏,并对动物健康产生负面影响。IEC 损伤通常与长期的急性应激有关,例如温度升高和新的应激管理模式。环境应激引起的肠上皮损伤与氧化应激有关。到目前为止,由于缺乏体外细胞培养系统,只能在体内检测饲料添加剂和处理对鸭肠道上皮抗氧化活性的影响。在这项研究中,我们描述了从小鸭胚胎小肠中简单分离和培养 IEC 的方案。免疫荧光用于 IEC 的细胞学鉴定。此外,还可以在培养细胞中检测到 IEC 标记基因(IAP 和 CDH1)。并进行细胞状态评估,通过 CCK-8 测定分析细胞增殖活力。此外,我们构建了氧化应激模型,用于研究氧化应激反应机制和作用于细胞信号转导途径的药物。总之,我们已经开发出一种有效且快速的方法来获得鸭原代 IEC,并构建了氧化应激模型。这些 IEC 表现出与上皮细胞一致的特征,可用于体外探索氧化应激的生理机制。